The assembly of actin filaments represents a dynamic biological process essential for an organism's survival. For example, actin filament dynamics provide the foundation for cellular motility necessary in the wound healing response and the neutralization of bacteria by neutrophils (Bamburg and Wiggin, 2002; Asch et al, 1996). Actin filament dynamics have further been identified as playing a key role in various pathologies including tumor metastasis, cystic fibrosis, organ infarction and glaucoma. It can be appreciated that, with the ever-increasing understanding of actin's integral role in disease and injury, there is a growing interest in identifying new and improved actin-targeted therapeutics.
Marine organisms, especially soft corals, sponges and tunicates, provide many secondary metabolites including the macrolides which are known to be cytotoxic through inhibition of actin filament dynamics (Saito et al, 1994; Wada et al, 1998; Bubb et al, 1995; Spector et al, 2002). These drugs can be classified as: (1), macrolides that contain a trisoxazole group and are typified by Kabiramide C (KabC; Matsunaga et al, 1986; Wada et al, 1998; FIG. 1A) and Ulapualide A (UlaA; FIG. 1A-C; Roesener and Scheuer, 1986); and (2), dimeric macrolides such as Swinholide A (SwiA; Bubb et al, 1995; Saito et al, 1996). A review of the chemical database details more than 30 different members within these macrolide families. Certain members of the trisoxazole macrolide family may act as actin monomer sequestering and severing drugs (Saito et al, 1996; Wada et al, 1998), while SwiA is thought to sequester actin dimers during the nucleation phase of filament growth (Bubb et al, 1995), and misakinolide A (MisA; Terry et al, 1997) is purported to bind to the actin dimer at the (+)-end of the filament where it blocks further growth. While understood to be effective in actin filament inhibition, the detailed molecular mechanism of how these natural products interact with actin was unknown before the inventor's present efforts, reported herein. Without this knowledge, the identification and rational design of macrolide functional derivatives could not be reliably carried out by those working in the field.
Based upon a need in the field, it is desirable to obtain additional macrolide analogs capable of, most preferably, unregulated F-actin (+)-end capping, F-actin severing and G actin monomer sequestration. The unregulated activity of such analogs would be in contrast to the regulated actin-capping proteins currently known such as gelsolin, CapG and Capping Protein (reviewed by Cooper and Schafer, 2000). The therapeutic use of these respective actin-capping proteins is hampered by their responsiveness to physiological cues such as second messengers including calcium and phosphatidyl inositol (e.g., PIP and PIP2,). Additional macrolide analogs would find use in, for example, the treatment of actin-containing clots, viscous mucus in cystic fibrosis, glaucoma, tumor growth and/or as a substitute for plasma gelsolin in patients suffering burns and trauma. Methods for screening and identifying new macrolide analogs based upon the present inventor's recent elucidation of macrolide:actin interactions would be particularly desirable and a welcome advancement in the field.